Obesity and its associated metabolic diseases are major health problems world-wide. Although the reasons for the rapid increase in rates of obesity are multifactorial, it is clear that neural circuits in the brain play a major role in sensing energy stores and regulating energy balance. These neural pathways are candidate targets for the development of new pharmacotherapies aimed at reversing the obesity epidemic and therefore it is essential that we understand their function in detail, including the complete map of interconnections with other neural systems and their utilization of multiple neurotransmitters and intracellular signaling pathways. This project focuses on a key component of the brain's energy balance circuitry, the proopiomelanocortin (POMC) neurons located in the hypothalamus and brainstem. Genetic deletion of POMC function from the brain results in profound obesity and a metabolic syndrome characterized by extreme hyperphagia and reduced basal metabolic rate. However, POMC neurons are heterogeneous in many aspects and accumulating evidence suggests that different subpopulations of the neurons regulate separate neurological processes that together result in normal or pathological control of caloric balance. The overall goals of this project are to identify specific functions of these neuronal subpopulations and the neuroanatomic and molecular pathways that they utilize. Specific aim 1 includes a series of behavioral and pharmacological studies to probe the underlying component processes and neural substrates contributing to hyperphagia in POMC-deficient mouse models. These experiments utilize a newly developed method for meal pattern analysis and will test the hypothesis that melanocortin signaling coordinately modulates stereotyped motor, reward, and hedonic aspects of feeding behavior; processes which are particularly relevant to human issues surrounding food choice and meal size in the clinical pathogenesis of obesity. Specific aim 2 will further define the neurocircuitry connecting arcuate POMC neurons to limbic forebrain nuclei. A novel retrograde tracing method involving site-specific microinjections of a canine adenoviral vector expressing Cre recombinase into mutant mice with a reversibly silenced POMC gene allele will be used to map axon collaterals to specific combinations of target sites. Second order neurons in limbic areas innervated by POMC terminals will be identified by trans-synaptic labeling with wheat germ agglutinin expressed in, and transported anterogradely from, POMC cell bodies. We will also use genetic techniques and multilabel immunohistochemistry to study the unexpectedly complex dendrites of POMC neurons that receive synaptic inputs from distal sites. Finally, in Specific aim 3 we will use complementary genetic approaches to study the unique functional role of spatially distinct POMC neuron subpopulations or developmentally altered POMC gene expression in the prevention or mitigation of obesity. The techniques involved are aggregation chimera formation and Cre recombinase-mediated reactivation of POMC expression from a neuron-specific and reversibly silenced POMC allele. PUBLIC HEALTH RELEVANCE: Among the greatest current threats to public health are the continually increasing rates of overweight, obesity, diabetes, and the metabolic syndrome. A complex set of neural circuits integrates the balance between caloric demand and utilization with the behavioral and psychological processes related to feeding. This project centers on a key component of the brain's feeding circuits, propiomelanocortin neurons, to explain how these neurons coordinately regulate appetite, meal initiation and termination, and metabolic rate to normally maintain body weight within tightly controlled limits.